A hollow fiber membrane for a purpose of liquid treatment has been widely utilized in industrial uses such as microfiltration and ultrafiltration and in medical uses such as hemodialysis and plasma separation. Particularly in recent years, there has been a demand in the field of pharmaceutical industry for an art of elimination of not only bacteria but also pathogenic substances in nano sizes such as virus during production steps so as to ensure high safety for bio-pharmaceutical drugs and blood products. The drug and the blood product as such are produced from a substance derived from living organisms such as protein by means of the steps such as incubation, recovery and purification. Accordingly, there is a risk of contamination of small amount of components derived from such materials and auxiliary materials (such as culture medium and water). Among them, virus contaminating therein is a highly risky component even if its amount is very small.
With regard to a method for removing and inactivating the virus, there are removal methods by means of a heating treatment, a highly energetic treatment with irradiation of gamma ray, ultraviolet ray, etc., a chemical treatment such as a treatment at low pH and a treatment using surfactant, a precipitation/fractionation method such as an ethanolic fractionation and an ammonium sulfate fractionation, chromatography and membrane filtration. Ability of a step for excluding the virus from the step is called a virus clearance. Among them, according to the membrane filtration method, affection to denaturation of protein to be recovered is small and even such a virus which is energetically and chemically resistant can be removed. Therefore, the membrane filtration method has been thought to be a useful method because it enables a sure separation/removal mainly due to a sieving effect. The membrane filtration method is a very reliable and highly efficient method in a process for removing virus from a solution of protein having smaller size than the virus so as to recover the protein. It is a matter of course that sharpness of separation size and completeness having no deficiency are demanded for a separation membrane used for removing the virus.
Moreover, in a process for producing bio-pharmaceutical drugs and blood products, protein which is a useful ingredient should be efficiently recovered in terms of productivity and yield. However, in such a case wherein the membrane has a pore size which guarantees sure removal of virus, sure removal of virus is guaranteed whereas property of protein recovery by permeation or life of the membrane lowers due to clogging of the membrane as the size of the protein to be recovered becomes near the size of virus. It is usual that, in such a use, pore size of the membrane is designed by placing a focus on the virus of the smallest size which is worried about as the contaminant virus (parvovirus having diameter of about 20 nm has been presumed to be a representative one). In that case, protein in an immunoglobulin region is a protein ingredient having the processible size in almost upper limit. Since the separation of protein and virus is conducted depending upon the sizes thereof as such, recovery of protein having bigger size than, for example, parvovirus is impossible by a membrane which guarantees removal of parvovirus. In such a case, only the virus clearance to virus in a size depending upon the membrane size (such as retrovirus) is guaranteed by a virus filtration membrane while, with regard to virus in small sizes permeating the membrane, the virus clearance of the step is guaranteed by a production process warranted by other removal or inactivating step. They are appropriately selected by producers depending upon the target protein of bio-pharmaceutical preparations and blood products to be recovered.
As mentioned above, when that which is predicted as the smallest virus in general is taken as a removal target, high removal of substances in a parvovirus size and good recovery of substances in a globulin size can be important indicators for the safety and the performance. In the process for producing drugs, it goes without saying that security of safety as quality of the preparations has the first priority. Therefore, it cannot be denied that the productivity such as permeation and recovery of protein becomes a victim to some extent. There has been a demand for the development of virus removing membrane which satisfies both of them. Since the productivity mentioned herein is also related to the cost of the preparations, there is a need for a production technique in a purifying step for providing the product at lower cost. Accordingly, it is the necessary technique in which the permeation characteristic of protein is enhanced while the pore size by which inhibition of virus can be surely inhibited is still retained.
Since a step for removing virus in bio-pharmaceutical drugs and blood products is carried out after a purifying step for achieving sufficient purity, there is such a characteristic that the factor affecting the permeability of the membrane to a solute is not the clogging and the blocking by a substance such as virus having bigger size than membrane pore size but the clogging due to the solute substance per se. Only a protein solution having such a property achieves the practical efficiency for the removal of virus by means of membrane separation. Accordingly, a step for filtrating almost pure protein solution can be said to be an actual virus removing step in a membrane method. In view of such a sense, it is likely that a decrease in permeability of a solute protein is caused by the clogging due to adsorption of protein per se (which can be said to have sufficiently high permeability in view of its size) with the pores and also by the blocking of pores.
It is supposed that adsorption of protein is mostly due to an interaction of the hydrophobic domain in protein and the hydrophobic surface of a membrane material. There is usually carried out a method wherein a hydrophilized membrane is used for a purpose of reducing the adsorption. This is a means which is widely used in membranes for blood purification, membranes for water purification, etc. as well. There have been conducted, for example, a method wherein membrane is manufactured from a hydrophilic polymer, a method wherein a hydrophobic polymer is a main constituent of a membrane and a hydrophilic polymer is blended in the materials to form a membrane, a method wherein, after a hydrophobic polymer membrane is manufactured, it is coated with a hydrophilic polymer.
Further, in order to achieve the higher productivity, it is also effective that not only the stability of filtration but also the permeation speed of the solution to be treated are made high. This is possible by making the water permeation coefficient of the membrane itself large and by making the operation pressure for the filtration high. Thus, the permeated liquid amount per unit time and unit area is determined by a product of the water permeation coefficient and the operation pressure. A designing matter for increasing the water permeation speed from the membrane structure is expansion of the pore size and reduction of the membrane thickness but, there is a limitation for warranting the exclusion of virus and for making the pore size large whereby that is not preferred. Accordingly, it is an effective membrane design that the membrane thickness is made as small as possible so that the resistance to water permeation is reduced. When the operation pressure is higher, the amount of permeated water can be increased but, since it also depends upon the durability of membrane and also upon the strength of piping of the apparatus as a whole, it is preferred to be made high within an extent allowable therefor. In a manufacturing step, it is usual that not only metal pipe but also silicon tube is used for the piping. Accordingly, an operation pressure is set in such a manner that its upper limit is about 3 to 4 bars. Desirable membrane is such a one which can be used as near as possible the upper limit within the above range. Needless to say, it is not preferred in view of guaranteeing the high virus-removing ability that application of pressure causes deformation of the membrane, changes and variations in the pore, etc. Accordingly, stability of the strong pore is also the necessary condition. Further, in such a case wherein a protein solution is filtrated at high pressure, there is a problem that, when adsorption to the membrane takes place, the adsorption becomes stronger due to the consolidation of a solute component whereby, for example, multi-layered adsorption is apt to be induced.
In addition, a very effective means for making the production of bio-pharmaceutical preparations and blood products efficient is to operate the liquid to be treated in a concentration of as high as possible. Although the concentration of the final preparation as a drug is decided in each of the products, in the intermediate processes for incubation and purification, treatment is not conducted as the liquid of this final concentration. When the treatment and the handling are conducted in a concentration of as high as possible, scale of the apparatus becomes compact and big advantages are resulted for the efficiency such as the time for feeding the liquid and the time for conducting the filtration operation. Accordingly, even in a filtration membrane, there is also a demand for the ability for treating a protein solution in a concentration of as high as possible within short time.
Patent Document 1 discloses a virus-retaining ultrafiltration membrane having a surface which is made hydrophilic using hydroxyalkyl cellulose. According to Patent Document 1, a surface of a hydrophobic polymer membrane is made hydrophilic using hydroxyalkyl cellulose, and the surface is treated in an autoclave or is immersed in boiling water whereby performance as a membrane for virus removal can be enhanced. The reason therefor is mentioned that hydrophilicity (angle of contact) increases by the treatment at 100° C. or higher and also that a hardly swelling state is resulted whereby improvement to a preferred mode is achieved. The resulting effect as such is likely to be an effect which is specific to the constitution of a membrane wherein a hydrophobic polymer is coated with hydroxyalkyl cellulose. It is not possible to further expand the effect to such a membrane wherein a membrane before the coating is hydrophilic (i.e. a membrane solely comprising a hydrophilic polymer or a blended membrane with a hydrophilic polymer). It is likely that, in the membrane as such, the membrane itself before the coating hardly retains the stability which can guarantee the inhibition of virus under the condition such as autoclaving or the membrane itself before the coating exhibits a swelling ability whereby it is presumed that the effect disclosed in this document cannot be achieved. In Patent Document 1, a filtrating operation at 30 psi (about 2 bar) is carried out and, as compared with a membrane wherein only a hydrophilic polymer is blended, it is likely that the stability of the operation at high pressure is also enhanced.
In a membrane wherein the hydrophilization of the membrane is done by a single hydrophilic polymer, it sometimes happens that a membrane surface wherein the hydrophilic polymer component is fully exposed is not always formed or all of the membrane surfaces or all of inner pore areas are not always coated when a hydrophobic polymer is blended with a hydrophilic polymer or is coated therewith. With regard to the causes therefor, it has been clarified according to the recent surface analysis technique that each of the hydrophobic and hydrophilic polymers is solely apt to have a domain structure and that, when the membrane surface is observed in a microscopic manner, those polymers (a hydrophilic polymer in many cases) are present in a separated state in a patch form. In that case, it is possible to further improve and optimize the hydrophilizing function by means of compounding a plurality of polymers. Moreover, the use of a hydrophilic polymer has a problem that elution of the hydrophilic polymer is resulted and there is a possibility that contamination of this component is generated in the treatment solution. Particularly in the case of use for medical purpose and for drug production, there is also predicted a sterilizing treatment by heat or by drug or a washing treatment and, in addition, a reuse or the like whereby its adaptability is important.
Further, when the membrane is coated with another hydrophilic polymer after formation of the membrane, it goes without saying that the possibility of the affection such as narrowing, clogging, etc. of the pores by the hydrophilic polymer used for the coating is to be taken into consideration. In that case, the relation between the molecular size of the polymer used for the coating and the pore size is particularly important and careful attention is necessary therefor. When a hydrophilic polymer having a sufficiently small molecular size to the pore size is used for the coating, risk of the clogging is relatively small but, when the molecular weight is small, no sufficient retention of adsorption ability is achieved and dropout is apt to happen.
In Patent Document 2, there is disclosed an art wherein a porous membrane consisting of a hydrophobic polymer is coated with a second hydrophilic polymer having higher hydrophilicity via a coating layer of a copolymer of polyvinyl alcohol with vinyl acetate. According to Patent Document 2, since the coating layer of the copolymer of polyvinyl alcohol with vinyl acetate is insoluble in water, it can form a stable coating layer for a hydrophobic polymer. On the other hand, since the copolymer of polyvinyl alcohol with vinyl acetate exhibits small hydrophilicity whereby it has no high suppressive ability for adsorption of protein or the like, the second highly hydrophilic polymer is further applied in order to improve such a point so as to achieve the hydrophilicity. A hydrophobic unit (vinyl acetate moiety) of the copolymer of polyvinyl alcohol with vinyl acetate contributes in enhancing the adsorption stability of the hydrophobic polymer of the substrate material while a hydrophilic unit (vinyl alcohol moiety) contributes in enhancing the adsorption stability of the second hydrophilic polymer. When the copolymer of polyvinyl alcohol with vinyl acetate is prepared by blending with a hydrophobic polymer, compatibility with the hydrophobic polymer is necessary but a high hydrogen-bonding ability of polyvinyl alcohol becomes a hindrance whereby it results in a phase separation from the hydrophobic polymer as well upon the stage of phase separation (resulting in a micro-domain structure) and, in addition, a solution composition in the preparation of a spinning dope is also very limited. Due to those reasons, it is presumed that those methods are substantially limited for the adaptation to a coating method as mentioned in Patent Document 2. In addition, when the second hydrophilic polymer is fixed to the first hydrophilic polymer (a copolymer of polyvinyl alcohol with vinyl acetate in Patent Document 2) by means of adsorption, it is desirable that the first hydrophilic polymer is not re-dissolved in a coating liquid in which the second hydrophilic polymer is dissolved. In Patent Document 2, there is disclosed such an idea, for example, to use a coating liquid in which saponification degree of the copolymer of polyvinyl alcohol with vinyl acetate is adjusted to low saponification degree exhibiting no water solubility. In addition, formation of a composite polymer thin membrane layer by means of a stepwise coating has such problems of the troublesomeness in membrane preparation by the treatment of coating solutions for a plurality of times and the affection to the substrate membrane itself by the influence of an organic solvent used for coating the copolymer of polyvinyl alcohol with vinyl acetate. Moreover, when an anchor effect to a hydrophobic polymer by the copolymer of polyvinyl alcohol with vinyl acetate is not fully achieved, there is also such a possibility that elution of a coating component becomes much and that hydrophilization becomes insufficient as well. When the hydrophobic solute (such as protein) contacts such a membrane, there is resulted a competitive adsorption with a coating polymer (in this case, it is a copolymer of polyvinyl alcohol with vinyl acetate) against a hydrophobic moiety of the substrate polymer and, if the adsorption with a solute component has a priority in terms of energy, exchange of the adsorbing component happens, which leads to the dropout of the coated polymer. In view of such a point, the polymer having a binder function contributing in a binding of a hydrophobic polymer with a highly hydrophilic polymer cannot be said to be sufficient only by means of the coating to a hydrophobic polymer. As such, Patent Document 2 uses the specificity by partial saponification of the copolymer of polyvinyl alcohol with vinyl acetate and it gives neither expansion to other materials and combinations nor suggestion therefor.
In Patent Document 3, there is shown a separation membrane consisting of (A) a hydrophobic polymer, (B) a polymer consisting of a hydrophilic unit only and (C) a copolymerized polymer consisting of a hydrophilic unit and a hydrophobic unit. To be more specific, (A) is a polysulfone type polymer, (B) is polyvinylpyrrolidone or polyethylene glycol and a hydrophilic unit in (C) is vinylpyrrolidone or ethylene glycol while a hydrophobic unit in (C) is vinyl acetate or vinylcaprolactam, and a membrane consisting of a blend of the component (A) and the component (B) is coated with the polymer of the component (C). In the constitution consisting of those three components, a membrane consisting of a hydrophobic polymer and a hydrophilic polymer (the component (A) and the component (B)) is further coated with the third component (the component (C)). When a copolymerized polymer partly having a hydrophobic unit is used instead of just a hydrophilic polymer as the third component, the hydrophobic unit exhibits an adsorbing ability to a hydrophobic moiety being exposed on the surface of a membrane consisting of a blend of two components which are a hydrophobic polymer and a hydrophilic polymer. As a result thereof, it is an object that the exposed hydrophobic moiety is mitigated by the component (C). Accordingly, it is ideal that surfaces of the membrane pores are formed only from the two components which are the hydrophobic polymer (the component (B)) and the copolymerized polymer (the component (C)). However, it can be easily presumed that the surface characteristic as such will result in a problem of extent that, when the affection by the exposed area of the component (A) is compared with the affection by the hydrophobic unit of the component (C), then which will be in less bad affection in terms of hydrophobicity. Thus, it is obvious that the membrane of the Patent Document 3 is inferior in terms of hydrophilicity to a membrane coated with the polymer which is entirely consisting of a hydrophilic unit (such as that in Patent Document 4). If and when a polymer solely consisting of a hydrophilic unit (such as cellulose type polymer, polyvinylpyrrolidone or polyethylene glycol which is exemplified in Patent Document 4) is used as the component (C) in Patent Document 3, the fixing force of the coat is weak and that is not preferred whereby there is a limitation in a combination of the component (B) with the component (C). Accordingly, in the art as shown in Patent Document 3, although the priority in a micron size such as adsorption of platelets mentioned in its text is noted, a high permeability of protein which is an object of the present invention cannot be expected. In addition, as mentioned already, the membrane before the coating (consisting of the component (A) and the component (B)) itself has a swelling property whereby it is predicted that the high stability of pore size and the resistance to pressure are not sufficient.